Fig. 4: Chronic stressed-induced senescence associated secretory phenotype (SASP) promotes senescence and fibrosis, as observed in FECD. | Cell Death & Disease

Fig. 4: Chronic stressed-induced senescence associated secretory phenotype (SASP) promotes senescence and fibrosis, as observed in FECD.

From: p16-mediated G0/G1 cell cycle arrest leads to SASP and fibrosis in Fuchs endothelial corneal dystrophy

Fig. 4: Chronic stressed-induced senescence associated secretory phenotype (SASP) promotes senescence and fibrosis, as observed in FECD.

a Schematic representation of the experimental pipeline used to evaluate the paracrine effects of SASP derived from acute (Acute SASP) and chronic stress conditions (Chronic SASP). b Brightfield images showing the morphology of untreated cells, Acute SASP-exposed cells with EMT-like features (white dotted circle), and chronic SASP-exposed cells with enlarged, flattened senescent-like morphology (white dotted circle). Scale bar: 50 µm. c SA-β-Gal staining (green) indicating an increased number of senescent cells exposed to chronic SASP. Scale: 25 µm. d Cell cycle analysis showing G0/G1 phase arrest in cells treated with acute and chronic SASP. e SA-β-Gal stain showing higher percentage of senescent cells after treating the tissues with acute and chronic SASP, indicative of senescent factors in the chronic SASP. Scale bar: 10 µm. f RT-PCR analysis showing relative gene expression of EMT (red), fibrosis (orange), ECM (green), senescence (blue) and FECD (purple) markers after acute and (g) chronic SASP, normalized to untreated controls of cadaveric human tissues. h Summary plot showing a higher fold increase in fibrotic, ECM and senescent markers observed after chronic SASP conditions. i multiplex cytokine array and ELISA results showing increased secretion of SASP factors including IL-6, MIP-1β, IFN-γ, TGFβIp from acute and chronic SASP. Chronic SASP additionally elevated IL-17, IL-8, MCP-1 and TNF-α levels exclusively. Data are presented as mean ± SEM; each dot represents one biological replicate. Statistical analysis: d two-way Anova with post-hoc Sidak’s multiple comparison test. c, e–i One-way Anova with post-hoc Tukey’s multiple comparison test. Statistical significance: ns=not significant; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. Biological replicates: n = 4 for all the analysis, and n = 3 for multiplex and ELISA analysis. Each donor tissue was divided in equal half.

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